Recent approval of anti-cancer immunotherapeutic drugs by the FDA validates that stimulation of the immune system of a cancer patient is an effective way to control or prevent metastatic disease. However, the approved immune checkpoint blockade inhibitors are effective only in a subset of melanoma patients, and a majority of the cancer patients do not respond to the drugs. Moreover, cancers such as breast cancers do not respond well to these drugs, suggesting additional immune stimulatory approaches such as therapeutic vaccination need to be developed. One of the major problems in developing vaccination-based immunotherapies for cancer is the heterogeneous nature of the disease. Recent deep gene sequencing studies demonstrated extensive intratumoral and interpersonal heterogeneity in gene mutations in tumors. Therefore, developing a therapeutic vaccine using a single antigen/peptide or patient-derived autologous/allogeneic tumor cell lines, which may not represent the heterogeneity of tumors, would not result in an immune response against the breadth of tumor antigens. Therefore, to induce immunity against the specific antigenic signature of a patient, use of an optimally adjuvanted vaccine based on whole autologous tumor tissue is highly desirable. We have developed an adjuvanted whole tumor tissue based vaccine for cancer immunotherapy using tumor membrane vesicles (TMVs) prepared from whole tumor tissue and modifying them by incorporating immunostimulatory molecules (ISMs) such as B7-1 and IL-12 using a novel protein transfer method. This method immobilizes ISMs onto TMVs via a glycolipid anchor (GPI), resulting in simultaneous delivery of TMVs along with biological adjuvants to APCs for enhanced uptake and optimal immune cell activation. Based on promising preclinical data in mice, Emory University and Metaclipse Therapeutics Corporation have entered into an agreement to advance this immunotherapy approach to clinics targeting triple negative breast cancer (TNBC). This subtype of cancer is highly heterogeneous and target antigens vary from patient to patient, thus making it an ideal target for our personalized immunotherapy. Metaclipse was awarded an NIH Phase I SBIR grant to further validate the technology in a TNBC model and is currently conducting studies required for IND filing. In this partnership grant application we propose to: 1) Perform preclinica studies to make recommendations on the use of the TMV-based, patient-specific immunotherapy in combination with standard-of-care treatments, 2) Investigate the effect of neoadjuvant chemotherapy on the yield and biological activity of the TMV-based immunotherapy prepared from mouse tumor tissue and human tumor tissue derived from patient-derived xenograft (PDX) models as well as TNBC patients and 3) Evaluate the TMV-based immunotherapy approach using human TNBC tumor tissue and human GPI-ISMs in a PDX model engrafted with a human immune system. The knowledge obtained from the proposed collaborative studies between Emory University and Metaclipse will aid in designing a clinical trial strategy for TMV-based immunotherapy in TNBC patients.